Wang Xiu-qing, Su Chun-xia, Zhou Bin, Cao Rui-bing, Chen Pu-yan
Key Laboratory of Animal Disease Diagnosis and Immunology, Ministry of Agriculture in Nanjing Agricultural University, Nanjing 210095, China.
Wei Sheng Wu Xue Bao. 2007 Feb;47(1):75-8.
According to the partiality codon of Pichia pastoris, hybrid antimicrobial peptide CecA-mag gene was synthesized and cloned into pPICZa-A to construct the recombinant expression vector pPICZa-A-CA. The SacI-linearized plasmid pPICZalpha-A-CA was transformed into P. pastoris SMD1168 by electroporation. Under the control of the promoter AOX'(alcoholoxidase') , an approximately 1.9kDa cecA-mag protein was expressed. Antibacterial assays demonstrated that cecA-mag had broad spectrum of antimicrobial property against Gram-positive as well as Gram-negative bacteria especially showed potent antibacterial activity against ampicillin resistant bacteria, such as pathogenic E. coli. In addition, the hybrid antibacterial peptide showed an extreme heat stable and acid stable characteristic. These results suggest that the P. pastoris expression system can be used to produce large quantities of fully functional cecA-mag for both research and industrial purpose. Based on these characteristics, the recombinant antibacterial peptide cecA-mag displays application foreground in the field of prevention of disease, and can be used as additives of animal feedstuff and so on.
根据毕赤酵母偏爱密码子，合成了杂合抗菌肽CecA-mag基因，并将其克隆到pPICZa-A中，构建重组表达载体pPICZa-A-CA。通过电穿孔将经SacI线性化的质粒pPICZalpha-A-CA转化到毕赤酵母SMD1168中。在启动子AOX'（醇氧化酶'）的控制下，表达了一种约1.9kDa的cecA-mag蛋白。抗菌试验表明，cecA-mag对革兰氏阳性菌和革兰氏阴性菌均具有广谱抗菌特性，尤其对氨苄青霉素耐药菌如致病性大肠杆菌表现出较强的抗菌活性。此外，该杂合抗菌肽具有极高的热稳定性和酸稳定性。这些结果表明，毕赤酵母表达系统可用于大量生产具有完全功能的cecA-mag，用于研究和工业目的。基于这些特性，重组抗菌肽cecA-mag在疾病预防领域具有应用前景，可作为动物饲料添加剂等。
